The banteng Bos javanicus is a globally threatened species of wild cattle restricted to South-East Asia. We report the first robust estimate of banteng density and population size from anywhere in the species' global range, using distance-based line transect sampling within two protected areas, Mondulkiri Protected Forest and Phnom Prich Wildlife Sanctuary, which form part of the Eastern Plains Landscape, Cambodia. We surveyed 110 line transects multiple times during the dry seasons of 2009–2010 and 2010–2011. In a total survey effort of 1,310 km there were 63 encounters with banteng. The mean estimate of the population across the 3,406 km2 study area is 3,200 (95% confidence interval 1,980–5,170). This suggests that the protected area complex of the Eastern Plains Landscape supports the majority of the global population of banteng. Stronger protection, both in the form of increased anti-hunting and poaching patrols and integrated land-use planning to prevent habitat loss within protected areas, is essential for securing wild cattle populations in the Eastern Plains Landscape.